The regulation of cytosolic calcium in rat brain synaptosomes by sodium-dependent calcium efflux.

1. When pinched-off presynaptic nerve endings (synaptosomes) isolated from rat brain are incubated in a low-Na (24-36 mM) medium, they take up 45Ca in a time-dependent manner. In a medium containing 1 mM-Ca, this Na-dependent 45Ca uptake amounts to approximately 10 nmol/mg protein at 1 min, and to approximately 40 nmol/mg protein at 20 min. The Na-dependent Ca uptake is not reduced when the synaptosomes are loaded with concentrations of quin 2 as high as 2 mM. 2. The increase in 45Ca uptake is paralleled by an increase in the free cytosolic Ca concentration [Ca]i, as monitored with the fluorescent Ca indicators quin 2 or fura 2. [Ca]i increases from the value of approximately 200 to approximately 500 nM within 3-5 min, and thereafter, remains at this elevated level. 3. When synaptosomes that have been loaded with 45Ca (for 1 min, in a low-Na medium) are diluted into an Na-containing medium, there is a rapid efflux of the Ca load. After correcting for Ca that is taken up during the efflux period, calculations show that the total Ca in the synaptosomes returns to the control level within 1 min. Measurements of total chemical Ca parallel the measurements made with radiotracer Ca, and confirm that the Ca loaded into the nerve terminals during a 5 min incubation in a low-Na medium is extruded from the nerve terminals within 1 min in a normal-Na medium. 4. The efflux of Ca from the synaptosomes is paralleled by a drop of [Ca]i to its basal level, also within 1 min. 5. The mitochondrial uncoupler, carbonyl cyanide p-trifluoromethyloxy-phenyl-hydrazone (FCCP, 1 microM), has no effect on either Na-dependent Ca uptake or efflux in synaptosomes. FCCP causes a slight (100-200 nM) increase in [Ca]i in synaptosomes resuspended in either a Na or a low-Na medium. This indicates that little of the Ca that is taken up by the synaptosomes in a low-Na medium is sequestered by the mitochondria. 6. These results suggest that Na-dependent Ca efflux (probably Na-Ca exchange) plays an important role in allowing nerve terminals to recover rapidly from a Ca load.